Abstract
A three-dimensional (3D) cell-based electrochemical sensor was developed to evaluate the immunomodulatory effects of atrazine and its metabolites by monitoring nitric oxide (NO) release. Nafion/Fe(III) meso-tetra (4-carboxyphenyl) porphyrin/reduced graphene oxide (Nafion/FeTCP@RGO) was functionalized on the screen-printed carbon electrode (SPCE) as a working electrode to enhance the NO selectivity and sensitivity. RAW264.7 cells encapsulated in a gelatin methacrylate (GelMA) hydrogel, forming a 3D cell culture system, were immobilized on the Nafion/FeTCP@RGO/SPCE to serve as biorecognition elements. The proposed sensor presented the induced NO expression as a distinct single differential pulse voltammetric (DPV) anodic peak in the selected physiological lipopolysaccharide (LPS) concentration range (0.01–2000 ng/mL). Furthermore, the peak current intensity of the NO was linearly correlated with the LPS concentration logarithm, which was further validated using a Griess reagent kit. The proposed sensor was utilized in optimized LPS-induction conditions to compare the immunosuppression effect of atrazine with its main metabolites (i.e. desethylatrazine (DEA), deisopropylatrazine (DIA), and diaminochlorotriazine (DACT)). The results demonstrated that the order of 30% inhibitory concentrations was atrazine (25.71 ± 1.08 μg/mL) < DEA (48.63 ± 2.17 μg/mL) ≤ DACT (49.11 ± 1.98 μg/mL) ≤ DIA (52.36 ± 2.34 μg/mL). Overall, this work provided a potential in vitro approach to evaluate the immunotoxicity of pesticides and their metabolites.
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